Extended Data Fig. 6: Analysis of the USP fold in the USP54~diUb-PA structure.
From: Discovery and mechanism of K63-linkage-directed deubiquitinase activity in USP53
a. Typical fold of USP DUBs in the structure of USP54~diUb-PA. The palm subdomain is colored red, the thumb subdomain blue and the fingers subdomain green. Additional structural elements including the blocking loops, the switching loop, the Cys loop, the zinc ions, and the catalytic triad are annotated. Close-up views for the coordination of all zinc ions are shown. b. Chains ABC, DEF, GHI and JKL of the crystal structure of USP54~diUb-PA, corresponding to the conformation in solution, are shown individually and as an overlay in cartoon representation. c. Gel-based cleavage assay assessing the catalytic cysteines. K63-linked tetraubiquitin chains (2 µM) were incubated with USP5320-383 (4 µM, upper) or USP5421-369 (300 nM, lower) and the respective catalytic cysteine mutants. Cleavage activity was analyzed by SDS-PAGE and Coomassie staining. d. Gel-based cleavage assay assessing the catalytic histidines. K63-linked triubiquitin chains (3 µM) were incubated with USP5320-383 (2 µM, upper) or USP5421-369 (300 nM, lower) and the respective catalytic histidine mutants for indicated time points. e. Gel-based cleavage assay as in d assessing potential oxy-anion hole residues. Uncropped versions of all gels are provided as Source Data.
